Molecular and pathological investigation of avian reovirus (ARV) in Egypt with the assessment of the genetic variability of field strains compared to vaccine strains

Avian orthoreovirus (ARV) is among the important viruses that cause drastic economic losses in the Egyptian poultry industry. Despite regular vaccination of breeder birds, a high prevalence of ARV infection in broilers has been noted in recent years. However, no reports have revealed the genetic and antigenic characteristics of Egyptian field ARV and vaccines used against it. Thus, this study was conducted to detect the molecular nature of emerging ARV strains in broiler chickens suffering from arthritis and tenosynovitis in comparison to vaccine strains. Synovial fluid samples (n = 400) were collected from 40 commercial broiler flocks in the Gharbia governorate, Egypt, and then pooled to obtain 40 samples, which were then used to screen ARV using reverse transcriptase polymerase chain reaction (RT-PCR) with the partial amplification of ARV sigma C gene. The obtained RT-PCR products were then sequenced, and their nucleotide and deduced amino acid sequences were analyzed together with other ARV field and vaccine strains from GenBank. RT-PCR successfully amplified the predicted 940 bp PCR products from all tested samples. The phylogenetic tree revealed that the analyzed ARV strains were clustered into six genotypic clusters and six protein clusters, with high antigenic diversity between the genotypic clusters. Surprisingly, our isolates were genetically different from vaccine strains, which aligned in genotypic cluster I/protein cluster I, while our strains were aligned in genotypic cluster V/protein cluster V. More importantly, our strains were highly divergent from vaccine strains used in Egypt, with 55.09–56.23% diversity. Sequence analysis using BioEdit software revealed high genetic and protein diversity between our isolates and vaccine strains (397/797 nucleotide substitutions and 148-149/265 amino acid substitutions). This high genetic diversity explains the vaccination failure and recurrent circulation of ARV in Egypt. The present data highlight the need to formulate a new effective vaccine from locally isolated ARV strains after a thorough screening of the molecular nature of circulating ARV in Egypt

Immunomodulating Effect of Echinacea and Star Anise in Protection and Treatment of Infectious Bronchitis Virus in Poultry

This study was carried out to evaluate the effect of two medicinal plants: Echinacea Purpura and Star anise for treating and prevention of infectious bronchitis virus (IBV) in chickens via assessment of their immune stimulating effect in IBV challenged chicks. 160 one day old Cobb® unsexed broiler chicks with average body weight 46.3 g. Chicks were classified into 8 equal groups (20 of each). The 1st group served as a control negative, the 2nd group served as a control positive (infected with IBV virus at day 21, non- treated), the 3rd group received Echinacea from the 7th till 21st day and infected with IBV virus at day 21, the 4th group received Star anise from the 7th till 21st day and infected with IBV virus at day 21, the 5th group received both Echinacea and Star anise from the 7th till 21st day and infected with IBV virus at day 21, the 6th group infected with IBV virus at day 21, then received Echinacea from the 21st till 42nd, the 7th group infected with IBV virus at day 21then received star anise from the 21st till 42nd, and the 8thgroup infected with IBV virus at day 21, then received both Echinacea and Star anise from the 21st till 42nd. Estimation of the collected samples (blood and sera) were made at different periods (7th, 14th, 28th and 35th days) to determine the effects of the used drugs on some hematological, and biochemical parameters. In addition, tissue specimens from liver and trachea were taken for histopathological examination. The obtained results evoked a significant increase in WBCs, heterophiles, lymphocytes, monocytes and esinophils counts in the groups treated with Echinacea and star anise compared with that of the control group. Serum ALT, AST, serum urea and creatinine results revealed a significant increase in groups treated with Echinacea and star anise compared with the control group, while GSH, SOD and NO revealed decrease in groups treated with Echinacea and star anise compared with the control group.  It could be concluded that the use of Echinacea and star anise as antivirals is positively beneficial in prevention and treatment of infectious bronchitis virus in poultry. Moreover, the use of the combination of both plants when used together have  more powerful effects in the prevention and treatment of IBV in poultry

Table_1_Molecular and pathological investigation of avian reovirus (ARV) in Egypt with the assessment of the genetic variability of field strains compared to vaccine strains.XLS

Avian orthoreovirus (ARV) is among the important viruses that cause drastic economic losses in the Egyptian poultry industry. Despite regular vaccination of breeder birds, a high prevalence of ARV infection in broilers has been noted in recent years. However, no reports have revealed the genetic and antigenic characteristics of Egyptian field ARV and vaccines used against it. Thus, this study was conducted to detect the molecular nature of emerging ARV strains in broiler chickens suffering from arthritis and tenosynovitis in comparison to vaccine strains. Synovial fluid samples (n = 400) were collected from 40 commercial broiler flocks in the Gharbia governorate, Egypt, and then pooled to obtain 40 samples, which were then used to screen ARV using reverse transcriptase polymerase chain reaction (RT-PCR) with the partial amplification of ARV sigma C gene. The obtained RT-PCR products were then sequenced, and their nucleotide and deduced amino acid sequences were analyzed together with other ARV field and vaccine strains from GenBank. RT-PCR successfully amplified the predicted 940 bp PCR products from all tested samples. The phylogenetic tree revealed that the analyzed ARV strains were clustered into six genotypic clusters and six protein clusters, with high antigenic diversity between the genotypic clusters. Surprisingly, our isolates were genetically different from vaccine strains, which aligned in genotypic cluster I/protein cluster I, while our strains were aligned in genotypic cluster V/protein cluster V. More importantly, our strains were highly divergent from vaccine strains used in Egypt, with 55.09–56.23% diversity. Sequence analysis using BioEdit software revealed high genetic and protein diversity between our isolates and vaccine strains (397/797 nucleotide substitutions and 148-149/265 amino acid substitutions). This high genetic diversity explains the vaccination failure and recurrent circulation of ARV in Egypt. The present data highlight the need to formulate a new effective vaccine from locally isolated ARV strains after a thorough screening of the molecular nature of circulating ARV in Egypt.</p

Molecular Detection of Reticuloendotheliosis Virus 5′ Long Terminal Repeat Integration in the Genome of Avipoxvirus Field Strains from Different Avian Species in Egypt

Avipoxviruses (APVs) are among the most complex viruses that infect a wide range of birds&rsquo; species. The infection by APVs is often associated with breathing and swallowing difficulties, reduced growth, decreased egg production, and high mortalities in domestic poultry. In the present study, 200 cutaneous nodular samples were collected from different avian species (chicken, pigeon, turkey, and canary) suspected to be infected with APVs from Dakahlia Governorate, Egypt. Pooled samples (n = 40) were prepared and inoculated in embryonated chicken eggs (ECEs). APVs were then identified by polymerase chain reaction (PCR) and sequence analysis of the APV P4b gene. Furthermore, the forty strains of APVs were screened for the presence of reticuloendotheliosis virus (REV)-5&prime;LTR in their genomes. Interestingly, the phylogenic tree of the APV P4b gene was separated into 2 clades: clade 1, in which our fowlpox virus (FWPV), turkeypox virus (TKPV), and canarypox virus (CNPV) isolates were grouped, along with reference FWPVs and TKPVs retrieved from GenBank, whereas, in clade2, the pigeonpox virus (PGPV) isolate was grouped with PGPVs retrieved from GenBank. Likewise, REV-5&prime;LTR was amplified from 30 strains isolated from chicken, turkey, and canary, while PGPV strains were free from REV-5&prime;LTR integration. To the best of our knowledge, this study involved the detection and characterization of REV-5&prime;LTR insertions in the APVs field isolates in Egypt for the first time. Given the above information, further future research seems recommended to understand the impact of the resulting REV-5&prime;LTR insertions on the pathogenesis, virulence, and inadequate vaccine protection against APVs

Molecular Characterization and Developing a Point-of-Need Molecular Test for Diagnosis of Bovine Papillomavirus (BPV) Type 1 in Cattle from Egypt

Bovine papillomatosis is a viral disease of cattle causing cutaneous warts. A diagnosis of this viral infection is very mandatory for combating the resulting economic losses. Given the limited data available about bovine papillomavirus (BPV) in Egypt, the present study involved the molecular diagnosis of bovine papillomavirus type-1 (BPV-1), -2, -4, -5, and -10 in cattle presenting cutaneous warts on the head and neck from New Valley Province, Egypt. The phylogenetic analysis of the detected types of BPV was also performed, followed by developing a point-of-need molecular assay for the rapid identification of identified BPV types. In this regard, a total of 308 cattle from private farms in Egypt were clinically examined, of which 13 animals presented cutaneous warts due to suspected BPV infection. The symptomatic animals were treated surgically, and biopsies from skin lesions were collected for BPV-1, -2, -4, -5, and -10 molecular identification using polymerase chain reaction (PCR). The presence of BPV-1 DNA was confirmed in 11 collected samples (84.6%), while BPV-2, -4, -5, and -10 were not detected. Sequencing of the PCR products suggested the Egyptian virus is closely related to BPV found in India. An isothermal nucleic acid amplification test (NAAT) with labeled primers specific for the BPV-1 L1 gene sequence, and based on recombinase polymerase amplification (RPA), in combination with a lateral flow strip assay for the detection of RPA products, was developed and tested. The point-of-need molecular assay demonstrated a diagnostic utility comparable to PCR-based testing. Taken together, the present study provides interesting molecular data related to the occurrence of BPV-1 in Egypt and reveals the genetic relatedness of the Egyptian BPV-1 with BPV-1 found in buffalo in India. In addition, a simple, low-cost combined test was also validated for diagnosis of the infection. The present study suggests the necessity of future investigations about the circulating strains of the virus among the cattle in Egypt to assess their genetic relatedness and better understand the epidemiological pattern of the disease

Molecular Characterization of Velogenic Newcastle Disease Virus (Sub-Genotype VII.1.1) from Wild Birds, with Assessment of Its Pathogenicity in Susceptible Chickens

Newcastle disease (ND) is considered to be one of the most economically significant avian viral diseases. It has a worldwide distribution and a continuous diversity of genotypes. Despite its limited zoonotic potential, Newcastle disease virus (NDV) outbreaks in Egypt occur frequently and result in serious economic losses in the poultry industry. In this study, we investigated and characterized NDV in wild cattle egrets and house sparrows. Fifty cattle egrets and fifty house sparrows were collected from the vicinity of chicken farms in Kafrelsheikh Governorate, Egypt, which has a history of NDV infection. Lung, spleen, and brain tissue samples were pooled from each bird and screened for NDV by real-time reverse transcriptase polymerase chain reaction (RRT-PCR) and reverse transcriptase polymerase chain reaction (RT-PCR) to amplify the 370 bp NDV F gene fragment. NDV was detected by RRT-PCR in 22 of 50 (44%) cattle egrets and 13 of 50 (26%) house sparrows, while the conventional RT-PCR detected NDV in 18 of 50 (36%) cattle egrets and 10 of 50 (20%) of house sparrows. Phylogenic analysis revealed that the NDV strains identified in the present study are closely related to other Egyptian class II, sub-genotype VII.1.1 NDV strains from GenBank, having 99.7–98.5% identity. The pathogenicity of the wild-bird-origin NDV sub-genotype VII.1.1 NDV strains were assessed by experimental inoculation of identified strains (KFS-Motobas-2, KFS-Elhamoul-1, and KFS-Elhamoul-3) in 28-day-old specific-pathogen-free (SPF) Cobb chickens. The clinical signs and post-mortem changes of velogenic NDV genotype VII (GVII) were observed in inoculated chickens 3 to 7 days post-inoculation, with 67.5–70% mortality rates. NDV was detected in all NDV-inoculated chickens by RRT-PCR and RT-PCR at 3, 7, and 10 days post-inoculation. The histopathological findings of the experimentally infected chickens showed marked pulmonary congestion and pneumonia associated with complete bronchial stenosis. The spleen showed histocytic cell proliferation with marked lymphoid depletion, while the brain had malacia and diffuse gliosis. These findings provide interesting data about the characterization of NDV in wild birds from Egypt and add to our understanding of their possible role in the transmission dynamics of the disease in Egypt. Further research is needed to explore the role of other species of wild birds in the epidemiology of this disease and to compare the strains circulating in wild birds with those found in poultry

Molecular Characterization of Velogenic Newcastle Disease Virus (Sub-Genotype VII.1.1) from Wild Birds, with Assessment of Its Pathogenicity in Susceptible Chickens

Newcastle disease (ND) is considered to be one of the most economically significant avian viral diseases. It has a worldwide distribution and a continuous diversity of genotypes. Despite its limited zoonotic potential, Newcastle disease virus (NDV) outbreaks in Egypt occur frequently and result in serious economic losses in the poultry industry. In this study, we investigated and characterized NDV in wild cattle egrets and house sparrows. Fifty cattle egrets and fifty house sparrows were collected from the vicinity of chicken farms in Kafrelsheikh Governorate, Egypt, which has a history of NDV infection. Lung, spleen, and brain tissue samples were pooled from each bird and screened for NDV by real-time reverse transcriptase polymerase chain reaction (RRT-PCR) and reverse transcriptase polymerase chain reaction (RT-PCR) to amplify the 370 bp NDV F gene fragment. NDV was detected by RRT-PCR in 22 of 50 (44%) cattle egrets and 13 of 50 (26%) house sparrows, while the conventional RT-PCR detected NDV in 18 of 50 (36%) cattle egrets and 10 of 50 (20%) of house sparrows. Phylogenic analysis revealed that the NDV strains identified in the present study are closely related to other Egyptian class II, sub-genotype VII.1.1 NDV strains from GenBank, having 99.7–98.5% identity. The pathogenicity of the wild-bird-origin NDV sub-genotype VII.1.1 NDV strains were assessed by experimental inoculation of identified strains (KFS-Motobas-2, KFS-Elhamoul-1, and KFS-Elhamoul-3) in 28-day-old specific-pathogen-free (SPF) Cobb chickens. The clinical signs and post-mortem changes of velogenic NDV genotype VII (GVII) were observed in inoculated chickens 3 to 7 days post-inoculation, with 67.5–70% mortality rates. NDV was detected in all NDV-inoculated chickens by RRT-PCR and RT-PCR at 3, 7, and 10 days post-inoculation. The histopathological findings of the experimentally infected chickens showed marked pulmonary congestion and pneumonia associated with complete bronchial stenosis. The spleen showed histocytic cell proliferation with marked lymphoid depletion, while the brain had malacia and diffuse gliosis. These findings provide interesting data about the characterization of NDV in wild birds from Egypt and add to our understanding of their possible role in the transmission dynamics of the disease in Egypt. Further research is needed to explore the role of other species of wild birds in the epidemiology of this disease and to compare the strains circulating in wild birds with those found in poultry

Consensus evidence-based recommendations for transition of care for adolescents with juvenile idiopathic arthritis: meeting patients’, parents’, and rheumatologists’ perspectives

Abstract Background Transition of care means the process of educating and empowering adolescents and young adults to take an active role in their own healthcare, develop decision-making skills, and eventually transition from paediatric to adult healthcare providers. Most people do not switch doctors until they are young adults, but it can be beneficial to start preparing children earlier. We aimed to develop a specific toolkit tailored to paediatric and adult rheumatologists to assist them in transitioning of care of young people with juvenile onset rheumatic musculoskeletal diseases from the paediatric to adult rheumatology care. Results The expert panel was confined to an online survey (n = 18), all the experts completed the two rounds. At the conclusion of round 2, a total of 10 points were gathered. The range of respondents (ranks 7–9) who agreed with the recommendations was 88.9 to 100%. All 10 clinical standards identified by the scientific committee were written in the same way. Based on the answers to the structured key questions and the literature review, a structured template was developed presenting transition of care integrated pathway. Conclusion The developed rheumatology-specific guideline offers adolescents and young adults a focussed, multidisciplinary transition of care approach with equity of access, quality of care and flexibility and set up standards for transitional care for young adults with juvenile rheumatological diseases

Global economic burden of unmet surgical need for appendicitis

Background There is a substantial gap in provision of adequate surgical care in many low- and middle-income countries. This study aimed to identify the economic burden of unmet surgical need for the common condition of appendicitis. Methods Data on the incidence of appendicitis from 170 countries and two different approaches were used to estimate numbers of patients who do not receive surgery: as a fixed proportion of the total unmet surgical need per country (approach 1); and based on country income status (approach 2). Indirect costs with current levels of access and local quality, and those if quality were at the standards of high-income countries, were estimated. A human capital approach was applied, focusing on the economic burden resulting from premature death and absenteeism. Results Excess mortality was 4185 per 100 000 cases of appendicitis using approach 1 and 3448 per 100 000 using approach 2. The economic burden of continuing current levels of access and local quality was US $92 492 million using approach 1 and$73 141 million using approach 2. The economic burden of not providing surgical care to the standards of high-income countries was $95 004 million using approach 1 and$75 666 million using approach 2. The largest share of these costs resulted from premature death (97.7 per cent) and lack of access (97.0 per cent) in contrast to lack of quality. Conclusion For a comparatively non-complex emergency condition such as appendicitis, increasing access to care should be prioritized. Although improving quality of care should not be neglected, increasing provision of care at current standards could reduce societal costs substantially

Global economic burden of unmet surgical need for appendicitis

Background There is a substantial gap in provision of adequate surgical care in many low- and middle-income countries. This study aimed to identify the economic burden of unmet surgical need for the common condition of appendicitis. Methods Data on the incidence of appendicitis from 170 countries and two different approaches were used to estimate numbers of patients who do not receive surgery: as a fixed proportion of the total unmet surgical need per country (approach 1); and based on country income status (approach 2). Indirect costs with current levels of access and local quality, and those if quality were at the standards of high-income countries, were estimated. A human capital approach was applied, focusing on the economic burden resulting from premature death and absenteeism. Results Excess mortality was 4185 per 100 000 cases of appendicitis using approach 1 and 3448 per 100 000 using approach 2. The economic burden of continuing current levels of access and local quality was US $92 492 million using approach 1 and$73 141 million using approach 2. The economic burden of not providing surgical care to the standards of high-income countries was $95 004 million using approach 1 and$75 666 million using approach 2. The largest share of these costs resulted from premature death (97.7 per cent) and lack of access (97.0 per cent) in contrast to lack of quality. Conclusion For a comparatively non-complex emergency condition such as appendicitis, increasing access to care should be prioritized. Although improving quality of care should not be neglected, increasing provision of care at current standards could reduce societal costs substantially